Early detection of Alzheimer’s
disease (AD) is
important
for taking proper measures against AD pathogenesis. Acetylcholinesterase
(AChE) is widely reported to be associated with the pathogenicity
of AD. Here, employing the “acetylcholine-mimic” approach,
we designed and synthesized a new class of naphthalimide (Naph)-based
fluorogenic probes for specific detection of AChE and avoiding interference
of butyrylcholinesterase (BuChE), the pseudocholinesterase. We investigated
the action of the probes on Electrophorus electricus AChE, and the native human brain AChE that we expressed in Escherichia coli and purified in the active form
for the first time. The probe Naph-3 exhibited a substantial
fluorescence enhancement with AChE and majorly avoided BuChE. Naph-3 successfully crossed the cell membrane of the Neuro-2a
cells and fluoresced upon reaction with endogenous AChE. We further
established that the probe could be effectively used for screening
AChE inhibitors. Our study provides a new avenue for the specific
detection of AChE, which can be extended to the diagnosis of AChE-related
complications.
Biosynthetically produced alkenes are high-value molecules that can serve as ‘drop-in’ replacements for fossil fuels. Alkenes are also heavily used in the polymer, lubricant, and detergent industries. UndB is the only known membrane-bound fatty acid decarboxylase that catalyzes the conversion of fatty acids to terminal alkenes at the highest reported in vivo titers. However, the enzyme remains poorly understood and enigmatic. Here, we demonstrate the first-time purification of UndB and establish that it is an oxygen-dependent, non-heme diiron enzyme that engages conserved histidine residues at the active site. We also identify redox partners that support the activity of UndB and determine the enzyme's substrate specificity and kinetic properties. We detect CO2 as the co-product of the UndB-catalyzed reaction and provide the first evidence in favor of the hydrogen atom transfer (HAT) mechanism of the enzyme. Our findings decipher the biochemistry of an enigmatic metalloenzyme that catalyzes 1-alkene biosynthesis at the membrane interface with the highest known efficiency.
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